Structural studies on P-TEFb and its role in regulation of transcription elongation

转录是基因转录成信使RNA的过程，信使RNA反过来又作为蛋白质合成的模板。转录的调节确保了细胞生长和分化所需的蛋白质的及时表达。转录调控中的错误可导致不受控制的细胞生长和增殖。正转录延伸因子B（P-TEF B）控制由RNA聚合酶II进行的转录的延伸阶段。P-TEFb不仅是绝大多数细胞基因转录所必需的，它还是人类免疫缺陷病毒HIV的关键宿主细胞辅因子。HIV使用P-TEFb使其自身基因组能够通过细胞转录装置转录。P-TEFb活性的增加也与其他疾病有关，例如心脏肥大和乳腺癌。我们的研究不仅旨在更好地了解P-TEFb如何识别其底物以及其活性如何在细胞内调节，而且还旨在确定P-TEFb特异性抑制剂的作用模式。一种这样的抑制剂是Flavopiridol，一种已经在临床试验中的潜在抗癌药物。这些研究将成为未来结构辅助药物发现计划的基础，以开发靶向P-TEFb的治疗药物。在细胞内，P-TEFb活性由其与分别称为BRD 4和HEXIM 1/7SK RNA的正调控因子和负调控因子的关联控制。我们的知识的机制，这些调节因子控制P-TEFb的活动将大大有助于了解其结构的分子细节。蛋白质结构可以用X射线晶体学和电子显微镜技术来确定。拟议的研究的目的是使用这两种技术来确定活性位点的分子细节以及大型抑制复合物的组织。这些信息将有助于我们进一步了解在转录延伸阶段转录活性是如何控制的，也将有助于我们了解HIV如何使用P-TEFb来促进其自身基因的转录。最终，我们可能能够利用这些知识来治疗疾病。

Transcription is the process during which genes are transcribed into messenger RNA that in turn serves as the template for protein synthesis. Regulation of transcription ensures the timely expression of proteins required for cell growth and differentiation. Errors in transcription regulation can lead to uncontrolled cell growth and proliferation. Positive transcription elongation factor b (P-TEFb) controls the elongation phase of transcription that is carried out by RNA polymerase II. Not only is P-TEFb essential for transcription of the vast majority of cellular genes, it is also a critical host cellular cofactor for the human immunodeficiency virus HIV. HIV uses P-TEFb to enable the transcription of its own genome by the cellular transcription apparatus. Increases in P-TEFb activity are also associated with other diseases, for example cardiac hypertrophy and breast cancer. Our study aims not only to contribute to a better understanding of how P-TEFb recognizes its substrates and how its activity is regulated within the cell, but also to identify the mode of action of specific inhibitors of P-TEFb. One such inhibitor is Flavopiridol, a potential drug against cancer that is already in clinical trials. These studies will form the basis for future structure-aided drug discovery initiatives to develop therapeutics targeting P-TEFb. Within the cell P-TEFb activity is controlled by its association with positive and negative regulatory factors, called BRD4 and HEXIM1/7SK RNA, respectively. Our knowledge of the mechanisms by which these regulatory factors control P-TEFb activity would be greatly assisted by knowing their structures in molecular detail. Protein structures can be determined using the techniques of X-ray crystallography and electron microscopy. The aim of the proposed research is to use both these techniques to characterise the molecular details of the active site as well as the organization of the large inhibitory complex. This information will be useful in furthering our understanding of how transcriptional activity is controlled at the stage of transcription elongation and will also facilitate our understanding of how HIV uses the P-TEFb to promote transcription of its own genes. Ultimately we may be able to exploit this knowledge for the treatment of disease.